During the current funding period two growth factor products of cardiogenic anterior endoderm, bone morphogenetic protein (BMP) and fibroblast growth factor (FGF), have been investigated with regard to how they act in concert to induce non-precardiac mesoderm to the cardiac lineage. Recently, findings in other laboratories have indicated that such combinatorial signaling may be integrated at the gene level by transcriptional co-activator proteins that contain histone acetyl transferase (HAT) domains. Our screen of a chick library yielded a HAT-containing protein that has been shown to be a member of the growing 'MYST' family. We recently demonstrated that this MYST is strongly and transiently expressed at the protein level in the developing chick, and mouse, myocardium; thus, this molecule has tentatively been termed 'cardiac' MYST (cMYST). During the renewal years we propose to investigate the role of cMYST in mouse heart development by performing four inter-related Aims. First (1) cMYST expression is being characterized in diseased and normal embryonic and adult tissues at the protein and mRNA levels. Concomitantly (2) we have characterized the genomic locus of cMYST and successfully targeted the cMYST gene in ES cells which are being used to prepare global cMYST mutants; we also plan to construct a cre/loxP targeting vectors to examine the effect of conditional ablation of the cMYST gene during mouse heart development. Third (3) the function of cMYST is being investigated via transfection assays using non-cardiac and cardiac cell-lines to determine whether cMYST is a transcriptional co-activator. This aim will also address the role of a cMYST isoform, as well as cMYST zinc finger, HAT and chromodomains, in transcriptional activation. Finally (4) it will be determined whether cMYST interacts with well-characterized cardiac transcription factors via immunoprecipitation and GST approaches, and/or whether it interacts with unknown novel factors to be identified using the yeast two-hybrid method. Elucidation of how these factors regulate cardiac myocyte differentiation will facilitate the design of approaches to repair the damaged or diseased myocardium in adults. This information is also relevant to understanding and ultimately managing congenital heart defects.